Activation of HIV-1 mRNA synthesis depends on the action of the viral trans-activator Tat, which exerts its effect on the processivity of viral transcription. Tat activity can be monitored by determining levels of promoter-proximal (nonprocessive) versus promoter distal (processive) transcription, which indicates the absence and presence of Tat function, respectively. We have demonstrated a predominance of nonprocessive transcription in a cell line model of latency (U1) and in PBMC from asymptomatic individuals, suggesting the occurrence of Tat-restricted transcriptional latency in vivo. Processive transcription can be detected following in vitro stimulation of these cells in culture. Three questions arise from these findings concerning viral behavior in vivo: to what extent does Tat-restricted transcription take place in vivo, do all or only a subset of CD4+ cells contain Tat-restricted virus and is this restriction reversible? The experiments of this proposal are designed to study the involvement of Tat in cellular latency and test the hypothesis that the regulation of Tat function is an important factor in the establishment and/or maintenance of clinical latency. These studies will use a novel RT-PCR assay for the specific detection of promoter-proximal and promoter-distal mRNA species in PBMC. With this assay we propose to: 1) Analyze patterns of transcriptional processivity in longitudinal samples of infected PBMC to evaluate the correlation with other parameters of infection and disease progression; cryopreserved archived specimens from the San Francisco Men's Health study and the Transfusion Safety Study will be accessed to address the prognostic potential of processivity determinations, 2) Study the inducibility of transcriptional processivity in latently-infected PBMC; these experiments will involve end-point dilutional analysis of mitogen activated PBMC to address questions concerning the reversibility of transcriptional latency and the proportion of PBMC that are transcriptionally latent, 3) Assess patterns of transcriptional processivity in T-cell subsets; these studies will characterize viral transcription within specific subpopulations of CD4+ T cells at different disease stages providing information on how compartmentalization of viral replication may contribute to immune dysfunction. Health relatedness: If discrete states of viral transcriptional activity can be defined with our assay, the course of disease and of therapy can be sensitively monitored long before other indicators of disease progression have changed.